【摘要】 将重组质粒pOTV-hSERT(oocytetranscriptionvector)中的人5-羟色胺转运体(hSERT)cDNA转录合成cRNA,通过显微注射技术注入爪蟾卵母细胞,建立hSERT异体表达模型,用电压钳技术测定其转运功能,研究hSERT重摄取的调控因素。胞外5-羟色胺(5-HT)灌流液引起内向转运电流,可被hSERT特异阻断剂去甲丙米嗪(desipramine)所阻断。该电流大小随测试电位的变化而改变,膜电位愈负,转运电流愈大。灌流液无钠或无氯时,转运电流分别减小(89.6±1.4)%和(51.7±1.5)%,表明胞外钠、氯离子为转运过程的必需条件。胞外5-HT浓度变化时,转运电流呈剂量依赖性,并有饱和现象。提高胞内5-HT浓度至胞外的20或40倍时,并不影响转运过程。因此,5-HT的重摄取与胞外Na+、Cl-离子联合转运有关,胞内5-HT浓度并不影响转运电流的大小,细胞膜电位的变化对转运过程有快速调控作用。更多还原

【Abstract】 cRNA of SERT was transcribed from cDNA, which was cloned in the pOTV vector. Oocytes of Xenopus laevis were injected with transcribed cRNA in vivo and turned to a heterologous expression model (human serotonin transporter, hSERT) in 5–6 days. With voltage clamp technique in recording the current induced by 5-HT, we tested the function of the expressed 5-HT transporter and studied factors which influence the re- uptake of hSERT. When the cell membrane potential was held at –60 mV and perfused with Ringer’s solution containing 5-HT, a steady inward current appeared and was almost completely inhibited by specific 5-HT trans- porter blocker, Desipramine, which demonstrated that it was 5-HT-induced current. The 5-HT-induced current varied with holding potentials or voltages in step protocol. The more negative of the clamping voltage, the bigger of the 5-HT-currents. The results from the experiments with 0 mmol/L Na+ or 0 mmol/L Cl– in Ringer solution indicated that transporting of 5-HT required Na+ and Cl– ion in extra-cellular solution and the influences of Na+ and Cl– ion on transporting were different, for with 0 mmol/L Na+ or 0 mmol/L Cl– the currents were reduced by (89.6±1.4)% and (51.7±1.5)% respectively. As the concentration of 5-HT in the perfusing Ringer’s solution increased from 0.1 to 15 μmol/L, the current augmented accordingly until it was saturated. Injection of 5-HT into the Oocyte making the intracellular concentration of 5-HT increased to about 20 or 40 folders, the 5-HT-induced current was not changed in spite of the changes of intracellular concentrations of 5-HT. Conclusion: Na+ ion and Cl– ion are co-transported with 5-HT. The cell membrane potential may influence the 5-HT-induced current fulfilling a rapid regulation for re- uptake of 5-HT. Changing the intracellular concentration of 5-HT does not influence the amplitude of transporter current.更多还原